Single-particle cryo-EM of the ryanodine receptor channel in an aqueous environment.

Ryanodine receptors (RyRs) are tetrameric ligand-gated Ca release channels that are responsible for the increase of cytosolic Ca concentration leading to muscle contraction. Our current understanding of RyR channel gating and regulation is greatly limited due to the lack of a high-resolution structure of the channel protein. The enormous size and unwieldy shape of Ca release channels make X-ray or NMR methods difficult to apply for high-resolution structural analysis of the full-length functional channel. Single-particle electron cryo-microscopy (cryo-EM) is one of the only effective techniques for the study of such a large integral membrane protein and its molecular interactions. Despite recent developments in cryo-EM technologies and break-through single-particle cryo-EM studies of ion channels, cryospecimen preparation, particularly the presence of detergent in the buffer, remains the main impediment to obtaining atomic-resolution structures of ion channels and a multitude of other integral membrane protein complexes. In this review we will discuss properties of several detergents that have been successfully utilized in cryo-EM studies of ion channels and the emergence of the detergent alternative amphipol to stabilize ion channels for structure-function characterization. Future structural studies of challenging specimen like ion channels are likely to be facilitated by cryo-EM amenable detergents or alternative surfactants.